Heavy alcohol use is common, yet avoidable cause of death, disability and human suffering. These adverse outcomes are potentially avoidable if heavy alcohol use is spotted early. However, current screening methods for chronic alcohol consumption capture alcohol usage only in the hours prior to testing or rely on insensitive, non-specific protein assays. In a recent highly successful Phase II project, using DNA from whole blood, we demonstrated that a combination of just two methylation sensitive droplet digital PCR (MSddPCR) assays could quite accurately identify subjects with recent heavy alcohol consumption. Since then, we have continued to advance those findings and have developed additional, even more powerful MSddPCR assays and refined our processes to only require the DNA found in a single drop of blood. In conjunction with our ethics and marketing team, we are now in the process of rolling this product nationwide. However, a significant barrier to a more universal implementation of this disruptive technology is the requirement for blood. Conceivably, a test that could use the DNA from saliva could circumvent patient concerns about the discomfort of a needle or lancet stick. In addition, it would enable both non-medically oriented alcohol treatment programs and telemedicine oriented treatment programs to more easily assess client response to treatment. This will be particularly critical for treatment of alcoholism in the elderly, who are often unable or unwilling to travel to alcohol treatment facilities. In this Phase I proposal, we intend to take the next steps to the creation of a saliva DNA screening test for heavy alcohol consumption. Already, we have collected and prepared saliva DNA on 150 subjects recently admitted to alcohol treatment and 150 age, gender and ethnicity matched controls. In addition, we have also developed an MSddPCR assay that can adjust for cellular heterogeneity in the saliva DNA. In this application, we will use these resources to whether MSddPCR assessments of saliva DNA can also be used to detect heavy alcohol consumption. Specifically, in Aim 1, we will determine methylation status at 4 CpG loci with high predictive power for detecting alcohol consumption and one locus whose information content can be used to correct for cellular admixture in saliva. Then, in Aim 2, we will conduct a series of quality assessment of the saliva DNA. Finally, in Aim 3, we will analyze the data from our 4 highly predictive loci, correcting when necessary for cellular admixture, to determine an algorithm capable of predicting heavy alcohol use status. In addition, we will analyze the quality control measures to identify those saliva DNA samples which give unreliable. As a result of this project, we will produce a set of MSddPCR assays and a quality control metric for a rigorous multisite Phase II test of this ?bloodless? NextGen product.